scholarly journals Establishment and Drug Screening of Patient-Derived Extrahepatic Biliary Tract Carcinoma Organoids

2021 ◽  
Author(s):  
Zhiwei Wang ◽  
Yinghao Guo ◽  
Yun Jin ◽  
Xiaoxiao Zhang ◽  
Hao Geng ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Biliary Tract ◽  
Drug Screening ◽  
Periodic Acid ◽  
Biliary Tract Carcinoma ◽  
Extrahepatic Cholangiocarcinoma ◽  
Whole Exome ◽  
Genetic Profiles

Abstract Background: Patient-derived organoids (PDO) have been proposed as a novel in vitro method of drug screening for different types of cancer. However, to date, extrahepatic biliary tract carcinoma (eBTC) PDOs have not yet been fully established. Methods: We collected six samples of gallbladder carcinoma (GBC) and one sample of extrahepatic cholangiocarcinoma (eCCA) from seven patients in order to attempt to establish eBTC PDOs for drug screening. Eventually, we successfully established five GBC PDOs and one eCCA PDO. Histological staining was used to compare the structural features of original tissues and cancer PDOs. Then, whole exome sequencing (WES) was used to analysed the genetic profiles of original tissues and cancer PDOs. Drug screening, including gemcitabine, 5-fluorouracil, cisplatin, paclitaxel, infigratinib, and ivosidenib, was measured and verified by some cases. Results: Different PDOs were found to have different growth rates during in vitro culture. Hematoxylin and eosin staining demonstrated that the structures of most cancer PDOs are able to retain the original structures of adenocarcinoma. Immunohistological staining and periodic acid-schiff staining were then discovered that marker expression in cancer PDOs were similar to those of the original specimens. Genetic profiles of the four original specimens, as well as paired cancer PDOs, were measured using whole exome sequencing. Three of the four PDOs exhibited a high degree of similarity, compared to the original specimens, except GBC2 PDO, which only had a concordance of 74% in the proportion of single nucleotide polymorphisms in the coding sequence. In general, gemcitabine was found to be the most efficient drug for treatments of the eBTCs, as it showed moderate or significant inhibitory impact on the growth of cancers. Results from the drug screening can be verified by three clinical cases, to a certain extent. Conclusions: Our study successfully established a series of eBTC PDOs, which helped fill in gaps in the field of eBTC PDOs. Additional measures should be explored to improve the growth rate of PDOs, and to preserve their immune microenvironment.

scholarly journals Establishment and drug screening of patient-derived extrahepatic biliary tract carcinoma organoids

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Zhiwei Wang ◽  
Yinghao Guo ◽  
Yun Jin ◽  
Xiaoxiao Zhang ◽  
Hao Geng ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Biliary Tract ◽  
Drug Screening ◽  
Periodic Acid ◽  
Biliary Tract Carcinoma ◽  
Clinical Effects ◽  
Whole Exome ◽  
Genetic Profiles

Abstract Background Patient-derived organoids (PDO) have been proposed as a novel in vitro method of drug screening for different types of cancer. However, to date, extrahepatic biliary tract carcinoma (eBTC) PDOs have not yet been fully established. Methods We collected six samples of gallbladder carcinoma (GBC) and one sample of extrahepatic cholangiocarcinoma (eCCA) from seven patients to attempt to establish eBTC PDOs for drug screening. We successfully established five GBC and one eCCA PDOs. Histological staining was used to compare structural features between the original tissues and cancer PDOs. Whole exome sequencing (WES) was performed to analyze the genetic profiles of original tissues and cancer PDOs. Drug screening, including gemcitabine, 5-fluorouracil, cisplatin, paclitaxel, infigratinib, and ivosidenib, was measured and verified by clinical effects in certain cases. Results Different PDOs exhibited diverse growth rates during in vitro culture. Hematoxylin and eosin staining demonstrated that the structures of most cancer PDOs retained the original structures of adenocarcinoma. Immunohistological and periodic acid-schiff staining revealed that marker expression in cancer PDOs was similar to that of the original specimens. Genetic profiles from the four original specimens, as well as paired cancer PDOs, were analyzed using whole exome sequencing. Three of the four PDOs exhibited a high degree of similarity when compared to the original specimens, except for GBC2 PDO, which only had a concordance of 74% in the proportion of single nucleotide polymorphisms in the coding sequence. In general, gemcitabine was found to be the most efficient drug for eBTC treatment, as it showed moderate or significant inhibitory impact on cancer growth. Results from drug screening were confirmed to a certain extent by three clinical cases. Conclusions Our study successfully established a series of eBTC PDOs, which contributed to the field of eBTC PDOs. Additional enhancements should be explored to improve the growth rate of PDOs and to preserve their immune microenvironment.

scholarly journals Whole-exome sequencing of cervical carcinomas identifies activating ERBB2 and PIK3CA mutations as targets for combination therapy

2019 ◽  
Vol 116 (45) ◽  
pp. 22730-22736 ◽  
Author(s):  
Luca Zammataro ◽  
Salvatore Lopez ◽  
Stefania Bellone ◽  
Francesca Pettinella ◽  
Elena Bonazzoli ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Cell Lines ◽  
Copy Number ◽  
Mtor Pathway ◽  
In Vivo Models ◽  
Whole Exome

The prognosis of advanced/recurrent cervical cancer patients remains poor. We analyzed 54 fresh-frozen and 15 primary cervical cancer cell lines, along with matched-normal DNA, by whole-exome sequencing (WES), most of which harboring Human-Papillomavirus-type-16/18. We found recurrent somatic missense mutations in 22 genes (including PIK3CA, ERBB2, and GNAS) and a widespread APOBEC cytidine deaminase mutagenesis pattern (TCW motif) in both adenocarcinoma (ACC) and squamous cell carcinomas (SCCs). Somatic copy number variants (CNVs) identified 12 copy number gains and 40 losses, occurring more often than expected by chance, with the most frequent events in pathways similar to those found from analysis of single nucleotide variants (SNVs), including the ERBB2/PI3K/AKT/mTOR, apoptosis, chromatin remodeling, and cell cycle. To validate specific SNVs as targets, we took advantage of primary cervical tumor cell lines and xenografts to preclinically evaluate the activity of pan-HER (afatinib and neratinib) and PIK3CA (copanlisib) inhibitors, alone and in combination, against tumors harboring alterations in the ERBB2/PI3K/AKT/mTOR pathway (71%). Tumors harboring ERBB2 (5.8%) domain mutations were significantly more sensitive to single agents afatinib or neratinib when compared to wild-type tumors in preclinical in vitro and in vivo models (P = 0.001). In contrast, pan-HER and PIK3CA inhibitors demonstrated limited in vitro activity and were only transiently effective in controlling in vivo growth of PIK3CA-mutated cervical cancer xenografts. Importantly, combinations of copanlisib and neratinib were highly synergistic, inducing long-lasting regression of tumors harboring alterations in the ERBB2/PI3K/AKT/mTOR pathway. These findings define the genetic landscape of cervical cancer, suggesting that a large subset of cervical tumors might benefit from existing ERBB2/PIK3CA/AKT/mTOR-targeted drugs.

Postmortem Whole Exome Sequencing Identifies Novel EIF2B3 Mutation With Prenatal Phenotype in 2 Siblings

2017 ◽  
Vol 32 (10) ◽  
pp. 867-870 ◽  
Author(s):  
Hannah Song ◽  
Sina Haeri ◽  
Hannes Vogel ◽  
Marjo van der Knaap ◽  
Keith Van Haren
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Clinical Care ◽  
Mutation Screening ◽  
Medical Decision ◽  
Homozygous Mutation ◽  
Vitro Fertilization ◽  
Whole Exome ◽  
The Impact

Objective: We describe 2 male siblings with a severe, prenatal phenotype of vanishing white matter disease and the impact of whole exome sequencing on their diagnosis and clinical care. Methods: The 2 children underwent detailed clinical characterization, through clinical and laboratory testing, as well as prenatal and postnatal imaging. Biobanked blood from the 2 siblings was submitted for whole exome sequencing at Baylor Laboratories. Results: Both male children had abnormal prenatal neuroimaging and suffered precipitous, fatal neurologic decline. Neuropathologic findings included subependymal pseudocysts, microcalcifications, and profound lack of brain myelin and sparing of peripheral nerve myelin. A novel homozygous mutation in the EIF2B3 gene (c.97A>G [p.Lys33Glu]) was found in both children; both parents were heterozygous carriers. The family subsequently conceived a healthy child via in vitro fertilization with preimplantation mutation screening. Conclusion: These histories expand the prenatal phenotype of eIF2b-related disorders and poignantly illustrate the impact that unbiased genomic sequencing can have on the diagnosis and medical decision making for families affected by childhood neurodegenerative disorders.

scholarly journals Digenic Variants in the FGF21 Signaling Pathway Associated with Severe Insulin Resistance and Pseudoacromegaly

2020 ◽  
Vol 4 (12) ◽  
Author(s):  
Stephen I Stone ◽  
Daniel J Wegner ◽  
Jennifer A Wambach ◽  
F Sessions Cole ◽  
Fumihiko Urano ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Hormone Secretion ◽  
Growth Hormone Secretion ◽  
Unknown Etiology ◽  
Fibroblast Growth Factor 21 ◽  
Severe Insulin Resistance ◽  
Whole Exome

Abstract Insulin-mediated pseudoacromegaly (IMPA) is a rare disease of unknown etiology. Here we report a 12-year-old female with acanthosis nigricans, hirsutism, and acromegalic features characteristic of IMPA. The subject was noted to have normal growth hormone secretion, with extremely elevated insulin levels. Studies were undertaken to determine a potential genetic etiology for IMPA. The proband and her family members underwent whole exome sequencing. Functional studies were undertaken to validate the pathogenicity of candidate variant alleles. Whole exome sequencing identified monoallelic, predicted deleterious variants in genes that mediate fibroblast growth factor 21 (FGF21) signaling, FGFR1 and KLB, which were inherited in trans from each parent. FGF21 has multiple metabolic functions but no known role in human insulin resistance syndromes. Analysis of the function of the FGFR1 and KLB variants in vitro showed greatly attenuated ERK phosphorylation in response to FGF21, but not FGF2, suggesting that these variants act synergistically to inhibit endocrine FGF21 signaling but not canonical FGF2 signaling. Therefore, digenic variants in FGFR1 and KLB provide a potential explanation for the subject’s severe insulin resistance and may represent a novel category of insulin resistance syndromes related to FGF21.

scholarly journals Η γενετική αρχιτεκτονική των εκ γενετής ανατομικών και νευρο-αναπτυξιακών διαταραχών

2021 ◽  
Author(s):  
Γεώργιος Κελλάρης
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Danio Rerio ◽  
Array Cgh ◽  
Loss Of Function ◽  
Whole Exome

Εισαγωγή: Οι νευρο-αναπτυξιακές ασθένειες, αποτελούν μια γενική κατηγορία ασθενειών οι οποίες σχετίζονταιμε την διαταραχή της ανάπτυξης και εύρυθμης λειτουργίας του εγκεφάλου και έχουν αρνητικές επιπτώσεις στηνοητική και συμπεριφορική λειτουργία των ασθενών. Οι ασθένειες αυτές εμφανίζουν μεγάλη κλινική και γενετικήετερογένεια καθώς περιλαμβάνουν συνδρομικές και μη συνδρομικές μορφές, συχνά δυσδιάκριτες που καθιστούνδύσκολη τη κλινική γενετική εκτίμηση και διάγνωση. Κατά συνέπεια, πολλές από αυτές τις περιπτώσειςπαραμένουν αδιάγνωστες και απαιτούν περαιτέρω διερεύνηση. Η αξιοποίηση των νέων τεχνολογιών όπως οΜοριακός Καρυότυπος (array-CGH) αλλά και η αλληλούχιση επόμενης γενιάς έχουν συμβάλει στην ανίχνευσηυποχρωμοσωμικών γενωμικών αναδιατάξεων και σημειακών παραλλαγών και στη ταυτοποίηση νέων γονιδίωνκαι παραλλαγών που μπορεί να συμβάλουν στο παρατηρούμενο φαινότυπο. Ταυτόχρονα, η αξιοποίησηπειραματικών ζωικών μοντέλων, όπως το zebrafish, έχει δειχθεί ότι μπορεί να συμβάλλει σημαντικά στηπροσπάθεια για τη κατανόηση των μοριακών και κυτταρικών διαδικασιών που σχετίζονται με την εμφάνισησυγκεκριμένων φαινοτύπων και το χαρακτηρισμό της παθογένειας των παραλλαγών που ανιχνεύονται στουςασθενείς.Σκοπός της μελέτης: Η παρούσα μελέτη επικεντρώνεται στη γενετική διερεύνηση ασθενών μενευροαναπτυξιακά και συγγενή νοσήματα, με αξιοποίηση των τεχνολογιών όπως ο Μοριακός Καρυότυπος(array-CGH) και η αλληλούχιση επόμενης γενιάς (Whole Exome Sequencing). Ταυτόχρονα, σε συνεργασία μεδιεθνώς αναγνωρισμένα Πανεπιστήμια του εξωτερικού αξιοποιήθηκε το πειραματικό ζωικό μοντέλο zebrafishγια τη μοντελοποίηση σπάνιων περιπτώσεων ασθενών με νευροαναπτυξιακά νοσήματα.Υλικό και μέθοδος: Υλικό του πρώτου μέρους της μελέτης, αποτέλεσε μια κοόρτη 97 ασθενών με σύνθετανευρο-αναπτυξιακά και συγγενή νοσήματα, όπως αλλαγές στο μέγεθος της κεφαλής (μικροκεφαλία, ήμακροκεφαλία), σε συνδυασμό με άλλα νευρο-αναπτυξιακά συμπτώματα. Για το πρώτο μέρος της μελέτηςαξιοποιήθηκαν o Μοριακός Καρυότυπος (array-CGH) και η αλληλούχιση επόμενης γενιάς (Whole ExomeSequencing). Υλικό του δεύτερου μέρους της μελέτης, αποτέλεσαν 2 αδέλφια αρσενικού γένους με νοητικήυστέρηση, μακροκεφαλία και πρόβλημα στην άρθρωση. Για το δεύτερο μέρος της μελέτης αξιοποιήθηκε τοπειραματικό ζωικό μοντέλο zebrafish για τη μοντελοποίηση των παραλλαγών που ανιχνεύθηκαν στους δύοασθενείς. Υλικό του τρίτου μέρους της μελέτης, αποτέλεσαν 7 ασθενείς από τρεις διαφορετικές οικογένειες,κλινικά χαρακτηριστικά όπως εντερική δυσλειτουργία και νευρολογικά συμπτώματα που παραπέμπουν σεδιάγνωση μιτοχονδριακής νευρο-εντερικής εγκεφαλο-μυοπάθειας. Για το τρίτο μέρος της μελέτης, αξιοποιήθηκαντο πειραματικό μοντέλο zebrafish και κυτταρολογικές μεθοδολογίες για τη μελέτη της λειτουργικότητας τωνμιτοχονδρίων για τη ταυτοποίηση ενός νέου υποψήφιου γονιδίου που σχετίζεται με την εμφάνιση τωνφαινοτύπων στους ασθενείς.Αποτελέσματα: Στο πρώτο μέρος της μελέτης, η χρωμοσωμική ανάλυση μικρο-συστοιχειών οδήγησε στηνανίχνευση παθολογικών και άγνωστης κλινικής σημασίας CNVs σε 31/75 άτομα που εξετάσθηκαν (41%). Απότο σύνολο των 45 CNVs που ανιχνεύθηκαν τα 29/45 (64%) αφορούσαν ελλείματα (deletions), τα 15/45 (33%)αφορούσαν διπλασιασμούς (duplications), ενώ παρατηρήθηκε και ένας τριπλασιασμός 1/45 (2%). Σε 13/75άτομα (17%) τα CNVs χαρακτηρίστηκαν ως παθογενή και σε 18/75 άτομα (24%) χαρακτηρίστηκαν ως άγνωστηςκλινικής σημασίας και χρήζουν περαιτέρω διερεύνησης. Ταυτόχρονα, η αλληλούχιση επόμενης γενιάς ανίχνευσε παθολογικές ή πιθανώς παθολογικές παραλλαγές σε γονίδια που σχετίζονται με την εμφάνιση κάποιου νευροαναπτυξιακού σύνδρομου, σε 39/95 άτομα (40.43%), ενώ σε 2/95 άτομα (2.13%) παρατηρήθηκανπαραλλαγές που άγνωστης κλινικής σημασίας σε γονίδια που προκαλούν κάποιο νευρο-αναπτυξιακό σύνδρομο. Από την άλλη πλευρά, σε 27/95 άτομα (28.7%) δεν ανιχνεύθηκε παραλλαγή σε κάποιο γονίδιο που έχει καταγραφεί ως τη προετοιμασία της παρούσας εργασίας να οδηγεί σε κάποιο γνωστό νευρο-αναπτυξιακό σύνδρομο και καταγράφηκαν ως αρνητικά (negative).Στο δεύτερο μέρος της μελέτης, αξιοποίηση της τεχνολογίας αλληλούχισης επόμενης γενιάς (Quad-basedwholeexome sequencing) επέτρεψε την ταυτοποίηση της μη-συνώνυμης παραλλαγής p.R79K στο γονίδιοDDX3X. Η παραλλαγή ανιχνεύθηκε στο δείγμα και των δύο ασθενών, ενώ η γενετική ανάλυση επιβεβαίωσε ότιη παραλλαγή κληρονομήθηκε από την ασυμπτωματική μητέρα (μητρικής προέλευσης), ενώ δεν ταυτοποιήθηκεκάποιο άλλο παθολογικό εύρημα. Στη συνέχεια, για να εξεταστεί η πιθανή συσχέτιση της παραλλαγή με τονφαινότυπο των ασθενών, αξιοποιήσαμε το πειραματικό μοντέλο danio rerio (zebrafish). Μελέτη τηςλειτουργικότητας της DDX3X με το zebrafish έδειξε ότι η παραλλαγή p.R79K οδηγεί σε απώλεια λειτουργίας(Loss of function) και είναι υπομορφική (hypomorphic).Στο τρίτο μέρος της μελέτης συμμετείχαν 7 ασθενείς από τρεις διαφορετικές οικογένειες, με κλινικά ευρήματαόπως εντερική δυσλειτουργία και νευρολογικά συμπτώματα που παραπέμπουν σε διάγνωση μιτοχονδριακήςνευρο-εντερικής εγκεφαλο-μυοπάθειας. Ανάλυση των κωδικών περιοχών του γενωμικού DNA με αλληλούχισηεπόμενης γενιάς (Whole exome sequencing), ανίχνευσε παραλλαγές σε ένα νέο γονίδιο, LIG3, το οποίο δεν έχειχαρακτηριστεί ως υπεύθυνο για την εμφάνιση κάποιας ασθένειας. Αξιοποίηση in vitro μεθοδολογιών με χρήσηκυττάρων απομονωμένα από ασθενείς έδειξε μειωμένα επίπεδα της πρωτεΐνης LIG3 ενώ μειωμένη ήταν και ηενεργότητα της λιγάσης. Παράλληλα, η δυσλειτουργία του LIG3 είχε αρνητική επίδραση στη συντήρηση τουμιτοχονδριακού DNA, με αποτέλεσμα την μείωση των επιπέδων μιτοχονδριακού DNA, χωρίς την παρουσίαπολλαπλών ελλείψεων που είναι χαρακτηριστικό άλλων μιτοχονδριακών ασθενειών. Έτσι, είναι πιθανό ηπαρατηρούμενη μιτοχονδριακή δυσλειτουργία να είναι υπεύθυνη για τη παρατηρούμενη παθολογία στουςασθενείς. In vivo λειτουργικές μελέτες με χρήση του πειραματικού μοντέλου danio rerio (zebrafish) έδειξαν ότι ηδυσλειτουργία του lig3 μπορεί να οδηγήσει σε παθολογικούς φαινοτύπους του εγκεφάλου και του εντέρου, σεαντιστοιχία με αυτούς που παρατηρήθηκαν στους ασθενείς.Συζήτηση/συμπεράσματα: Η αξιοποίηση των σύγχρονων τεχνολογιών όπως o Μοριακός Καρυότυπος και ηαληλούχιση επόμενης γενιάς (Whole Exome Sequencing) επιτρέπουν τη γρήγορη και ακριβέστερή διάγνωση καιτη καλύτερη συσχέτιση φαινοτύπου/γονοτύπου και κατά συνέπεια την ορθότερη και πιο ολοκληρωμένη γενετικήσυμβουλευτική. Όταν είναι δυνατό, η αξιοποίηση in vitro μεθοδολογιών για τη περαιτέρω διερεύνηση τωνμοριακών μηχανισμών που σχετίζονται με το παρατηρούμενο φαινότυπο αλλά και πειραματικών ζωικώνμοντέλων όπως το zebrafish μπορεί να συμβάλλει στη ταυτοποίηση νέων γονιδίων και το χαρακτηρισμό τωνπαραλλαγών που ανιχνεύονται σε αυτά.

scholarly journals Adrenal Insufficiency, Sex Reversal, and Angelman Syndrome due to Uniparental Disomy Unmasking a Mutation in CYP11A1

2018 ◽  
Vol 89 (3) ◽  
pp. 205-210 ◽  
Author(s):  
Ahlee Kim ◽  
Masanobu Fujimoto ◽  
Vivian Hwa ◽  
Philippe Backeljauw ◽  
Andrew Dauber
Keyword(s):  
Adrenal Insufficiency ◽  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Angelman Syndrome ◽  
Uniparental Disomy ◽  
In Vitro Studies ◽  
Recessive Mutation ◽  
Primary Adrenal Insufficiency ◽  
Whole Exome

Background/Aims: Cholesterol side-chain cleavage enzyme (P450scc) deficiency is a rare genetic disorder causing primary adrenal insufficiency with or without a 46,XY disorder of sexual development (DSD). Herein, we report a case of the combination of primary adrenal insufficiency, a DSD (testes with female external genitalia in a setting of a 47,XXY karyotype), and Angelman syndrome. Methods: Comprehensive genetic analyses were performed, including a single nucleotide polymorphism microarray and whole-exome sequencing. In vitro studies were performed to evaluate the pathogenicity of the novel mutation that was identified by whole-exome sequencing. Results: The patient was found to have segmental uniparental disomy (UPD) of chromosome 15 explaining her diagnosis of Angelman syndrome. Whole-exome sequencing further revealed a novel homozygous intronic variant in CYP11A1, the gene encoding P450scc, found within the region of UPD. In vitro studies confirmed that this variant led to decreased efficiency of CYP11A1 splicing. Conclusion: We report the first case of the combination of 2 rare genetic disorders, Angelman syndrome, and P450scc deficiency. After 20 years of diagnostic efforts, significant advances in genetic diagnostic technology allowed us to determine that these 2 disorders originate from a unified genetic etiology, segmental UPD unmasking a novel recessive mutation in CYP11A1.

scholarly journals TMOD-31. NOVEL GENETIC CHARACTERIZATION OF MURINE MEDULLOBLASTOMA CELL LINES

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii234-ii234
Author(s):  
Sarah Rumler ◽  
Mark Maienschein-Cline ◽  
Melline Fontes Noronha ◽  
Fei Song ◽  
Andrew Lock ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Cell Lines ◽  
Genetically Engineered ◽  
Myc Oncogene ◽  
Dna And Rna ◽  
Whole Exome ◽  
Genetically Engineered Mice ◽  
Genome Level

Abstract Pre-clinical research in medulloblastoma (MB) is conducted either with human-derived cell lines or tumors arising in genetically engineered mice (GEM). Murine MB cell lines are not readily available. To begin in vitro immuno-oncology MB research, we identified murine MB cell lines from research laboratories to determine their fidelity to human cell lines and GEM models. Using PubMed, we identified murine MB cell lines of potential interest. Seven cell lines form two independent laboratories were identified. We characterized the cell lines’ behavior in culture and performed next generation sequencing (NGS) on four of them (two from each group) to determine the extent to which these cell lines resemble both human and GEM MB models. DNA and RNA were extracted from MB cell lines, quality and quantity assessed. Whole-exome sequencing was performed and post-capture libraries sequenced. The cell lines identified were derived from either FVB/N TP53 null mice carrying the N-MYC oncogene (GTML3 and GTML5) or C57Bl6/J TP53 null heterozygous ptch (-/+) mice (MM1 and MM3). GTML cells grow in suspension and form clusters. MM cells are adherent, growing in monolayers. Whole exome sequencing of the 4 lines revealed similarities between both GEM and human MB cell lines, demonstrating 25% of sixteen genes known to be affected in both GEM tumors and human MB cell lines. Furthermore, the four cell Iines shared five deleterious mutations previously not described in MB, suggesting they might play a role in MB tumorigenesis: Pde4d, Whrln, Olr156, RIKEN cDNA D630045J12, and Olfr159. These are the first data to describe these murine MB cell lines and demonstrates 1) they resemble both GEM tumors and human MB cell lines at the genome level, and 2) they offer the potential to identify heretofore unrecognized mechanisms in MB tumorigenesis.

scholarly journals Somatic Mutations from Whole Exome Sequencing Analysis of the Patients with Biliary Tract Cancer

2018 ◽  
Vol 16 (4) ◽  
pp. e35
Author(s):  
Kyong-Ah Yoon ◽  
Sang Myung Woo ◽  
Yun-Hee Kim ◽  
Sun-Young Kong ◽  
Sung-Sik Han ◽  
...  
Keyword(s):  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Biliary Tract ◽  
Biliary Tract Cancer ◽  
Somatic Mutations ◽  
Sequencing Analysis ◽  
Tract Cancer ◽  
Whole Exome

scholarly journals Host Genetics and Antiviral Immune Responses in Adult Patients With Multisystem Inflammatory Syndrome

2021 ◽  
Vol 12 ◽  
Author(s):  
Andreas Ronit ◽  
Sofie E. Jørgensen ◽  
Casper Roed ◽  
Robert Eriksson ◽  
Ulrik W. Iepsen ◽  
...  
Keyword(s):  
Immune Responses ◽  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Mononuclear Cells ◽  
Type I ◽  
Sequencing Analysis ◽  
Healthy Controls ◽  
Whole Exome ◽  
Inflammatory Syndrome

COVID-19 associated multisystem inflammatory syndrome (MIS) is a rare condition mostly affecting children but also adults (MIS-A). Although severe systemic inflammation and multiorgan dysfunction are hallmarks of the syndrome, the underlying pathogenesis is unclear. We aimed to provide novel immunological and genetic descriptions of MIS-A patients. Cytokine responses (IL-6, IL-1β, TNFα, CXCL10, type I, II and III interferons) following SARS-CoV-2 infection of peripheral blood mononuclear cells in vitro were analyzed as well as antibodies against IFNα and IFNω (by ELISA) in patients and healthy controls. We also performed whole exome sequencing (WES) of patient DNA. A total of five patients (ages 19, 23, 33, 38, 50 years) were included. The patients shared characteristic features, although organ involvement and the time course of disease varied slightly. SARS-CoV-2 in vitro infection of patient PBMCs revealed impaired type I and III interferon responses and reduced CXCL10 expression, whereas production of proinflammatory cytokines were less affected, compared to healthy controls. Presence of interferon autoantibodies was not detected. Whole exome sequencing analysis of patient DNA revealed 12 rare potentially disease-causing variants in genes related to autophagy, classical Kawasaki disease, restriction factors and immune responses. In conclusion, we observed an impaired production of type I and III interferons in response to SARS-CoV-2 infection and detected several rare potentially disease-causing gene variants potentially contributing to MIS-A.

Genomic ERBB2/ERBB3 mutations promote PD-L1-mediated immune escape in gallbladder cancer: a whole-exome sequencing analysis

Gut ◽  
2018 ◽  
Vol 68 (6) ◽  
pp. 1024-1033 ◽  
Author(s):  
Maolan Li ◽  
Fatao Liu ◽  
Fei Zhang ◽  
Weiping Zhou ◽  
Xiaoqing Jiang ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Exome Sequencing ◽  
Whole Exome Sequencing ◽  
Ectopic Expression ◽  
Sequencing Analysis ◽  
Antibody Treatment ◽  
Whole Exome ◽  
Recurrent Mutations

ObjectivesPatients with gallbladder carcinoma (GBC) lack effective treatment methods largely due to the inadequacy of both molecular characterisation and potential therapeutic targets. We previously uncovered a spectrum of genomic alterations and identified recurrent mutations in the ErbB pathway in GBC. Here, we aimed to study recurrent mutations of genes and pathways in a larger cohort of patients with GBC and investigate the potential mechanisms and clinical significance of these mutations.DesignWe performed whole-exome sequencing (WES) in 157 patients with GBC. Functional experiments were applied in GBC cell lines to explore the oncogenic roles of ERBB2/ERBB3 hotspot mutations, their correlation with PD-L1 expression and the underlying mechanisms. ERBB inhibitors and a PD-L1 blocker were used to evaluate the anticancer activities in co-culture systems in vitro and in vivo.ResultsWES identified ERBB2 and ERBB3 mutations at a frequency of 7%–8% in the expanded cohort, and patients with ERBB2/ERBB3 mutations exhibited poorer prognoses. A set of in vitro and in vivo experiments revealed increased proliferation/migration on ERBB2/ERBB3 mutation. Ectopic expression of ERBB2/ERBB3 mutants upregulated PD-L1 expression in GBC cells, effectively suppressed normal T-cell-mediated cytotoxicity in vitro through activation of the PI3K/Akt signalling pathway and contributed to the growth and progression of GBC in vivo. Treatment with an ERBB2/ERBB3 inhibitor or a PD-L1 monoclonal antibody reversed these immunosuppressive effects, and combined therapy revealed promising therapeutic activities.ConclusionsERBB2/ERBB3 mutations may serve as useful biomarkers in identifying patients who are sensitive to ERBB2/ERBB3 inhibitors and PD-L1 monoclonal antibody treatment.Trial registration numberNCT02442414;Pre-results.

Sign in / Sign up

Export Citation Format

Share Document